As a molecular switch the ras protein undergoes structural changes that couple recognition sites on the surface of the protein to the guanine nucleotide-divalent metal ion binding site. X-ray crystallographic studies of p2l suggest that coordination between threonine-35 and the divalent metal ion plays an important role in these conformational changes. Recent ESEEM studies of p2l in solution, however, place threonine-35 further away from the metal and were interpreted as weak or indirect coordination of this residue. We have reported the high frequency (139.5 GHz) EPR spectra of p2l*Mn(II) complexes of two alternate guanine nucleotides that probe the link between threonine-35 and the divalent metal ion. In particular, the number of water molecules in the first coordination sphere [unreadable]of the manganous ion was determined to be four in p2 1 -Mn-(Il)-GDP and two in p2 I -Mn(H)-GMPPNP. The results for GMPPNP (a GTP-analog) are consistent with the number of water molecules predicted by the X-ray structure. These results rule out indirect coordination of threonine-35 and are consistent with direct, weak coordination of this residue as suggested by Halkides. The 170 hyperfine coupling constant of H2 170 is determined as 0.25 mT in the GDP form and 0.28 mT in the GTP form. These values are similar to reported values for 170-enriched aquo- and phosphato-ligands in other complexes of Mn(II). For all of these measurements, the high magnetic field strength (4.9 T), corresponding to 139.5 GHz EPR excitation, yields narrow Mn(II) linewidths and thus enhances sensitivity to 170 hyperfine broadening.